R: qPCR Experiment for the Amplification of HPRT1 Using the...
C126EG595
R Documentation
qPCR Experiment for the Amplification of HPRT1 Using the Bio-Rad iQ5 thermo cycler
Description
A quantitative PCR (qPCR) with the DNA binding dye (EvaGreen) (Mao et al. 2007)
was performed in a Bio-Rad iQ5 thermo cycler. The cycle-dependent increase
of the fluorescence was quantified at the annealing step (59.5 degrees Celsius).
Usage
data(C126EG595)
Format
A data frame with 40 observations on the following 97 variables. The first
column ("Cycle") contains the number of cycles and consecutive columns
contain the replicates ("A01" to "H12").
Details
HPRT1 was amplified in the Bio-Rad iQ5. The the change of
fluorescence was simultaneously monitored for the Hydrolysis probe of HPRT1
and EvaGreen. The primer sequences for HPRT1 were taken from Roediger et al.
(2013). A 10 micro L qPCR reaction was composed of 250 nM primer (forward and
reverse), qPCR Mix (according to the manufactures recommendations), 1 micro L
template (HPRT1 amplification product), 60 nM hydrolysis probe probe for
HPRT1. EvaGreen was used at 0.5 x final. During the amplification was monitored
59.5 degrees Celsius.
Source
Stefan Roediger, Claudia Deutschmann (BTU Cottbus - Senftenberg)
References
A Highly Versatile Microscope Imaging Technology Platform for the Multiplex
Real-Time Detection of Biomolecules and Autoimmune Antibodies. S. Roediger,
P. Schierack, A. Boehm, J. Nitschke, I. Berger, U. Froemmel, C. Schmidt,
M. Ruhland, I. Schimke, D. Roggenbuck, W. Lehmann and C. Schroeder.
Advances in Biochemical Bioengineering/Biotechnology. 133:33–74, 2013.
http://www.ncbi.nlm.nih.gov/pubmed/22437246
Mao, F., Leung, W.-Y., Xin, X., 2007. Characterization of EvaGreen and the
implication of its physicochemical properties for qPCR applications.
BMC Biotechnol. 7, 76.